The aim of this study was to assess the effects of CAP on captive Iberian lynx males, and evaluate its potential role in reducing short and long-term stereotypies and other abnormal behaviours. Regarding the latter, the test group showed a significant decrease in the time spent performing stereotypic behaviours between the baseline period (11.24%) and the CAP treatment period (3.68%) (Figure 3). These results concur with those of Spielman (2000) who found pheromones to reduce pacing in captive lions and tigers. Spielman also reported that after the completion of treatment, when the application of pheromones no longer took place, the pacing increased. Sajjad et al. (2011) also found that pacing increased for tigers held in a non-enriched environment. Macri & Patterson-Kane (2011) reported that pacing slightly decreased for solitary housed snow-leopards, although it increased for socially housed animals, when both groups were treated with synthetic analogues of cat pheromones.

The test group in the present study showed no significant differences between the time spent performing Relaxed behaviours during the baseline and the treatment periods, spending only about 5% of their time performing such behaviours. The control group spent more time performing these behaviours, although there was a non-significant decrease between the baseline’s 41% and the treatment’s 30% (Figure 11). This contrasts with the findings of Pitsko (2003) that found tigers to spend 76% of their time resting. The present study’s results also contrast with Macri & Patterson-Kane (2011) and Martínez-Macipe et al. (2015) who reported snow-leopards and lions to spend 53% and 92% of their time, respectively, performing relaxed behaviours. The differences seen between these studies and the present one might be due to the time of the day when sessions took place. Bashaw et al. (2007) showed that large felids are active mostly at dawn and dusk and spend most time resting around midday. Penadab et al. (2012) argued the same and also found that the captive Iberian lynxes in the El Acebuche centre were mostly active between 9:00 and 13:00, which coincided with the time when keepers were around the enclosures, doing the morning routines. This daily routine is similar to the one at the CNRLI (both centres are part of the same Breeding Program), and the observations of the present study were conducted in the morning, usually between 8:00 and 12:00. This might explain the low percentage of time spent performing Relaxed behaviours by the lynxes used in this study, compared to other captive wild felids.

Feliway® and CAP have been shown to reduce urine marking in domestic cats (Pageat&Tessier, 1997; Frank & Houpt, 1999; DePorter, 2013), however, the results of Macri & Patterson-Kane (2011) and of the present study contradict that. Urine marking was categorised as a Territorial behaviour, as some authors consider scent marking and scratching to be territorial marking behaviours (Pageat et al., 2010). The test group performed these behaviours in 28% of the observed time in the baseline period, increasing to about 43% during the CAP treatment period (Figure 15).

Just like Gaultier et al. (2005) found in their study with tigers, lynxes also share behavioural traits with domestic cats, such as urine- and paw-marking, and grooming. In addition, both species show obvious individual personality differences. Some of the techniques used to prevent undesired behaviours in domestic cats can be applied to wild felids in captivity (Gaultier et al., 2005), including pheromone therapy. To the best of my knowledge, there are no synthetic analogues of wild felids’ pheromones, for that reason a synthetic analogue of a domestic cat pheromone was used in the present study. Lynxes and domestic cats are part of the Felidae family and their semiochemicals have been shown to be effective on other species within this family, when the aim was to reduce abnormal behaviours and/or increase species-specific ones (Spielman, 2000; Gaultier et al., 2005; Macri & Patterson-Kane, 2011; Martínez-Macipe et al., 2015). Even though most of the mentioned studies used Feliway®, it has been shown that CAP also has positive effects. Cozzi et al., (2010) found that CAP increased affiliative interactions between domestic cats and Martínez-Macipe et al. (2015) reported that it increased social and play activity in captive lions. No similar conclusions can be drawn from the present study, since the animals were not housed in groups. However, the used ethogram included play-like behaviours that, despite not being observed, would have been included in the Exploratory behaviours category. The control group spent more time performing such behaviours (39% in both periods) than the test group. The latter group showed a non-significant decrease in the time spent performing Exploratory behaviours between the baseline period (27%) and the CAP treatment period (21%) (Figure 16). These results are not in agreement with the previously mentioned findings by Cozzi et al., (2010) and Martínez-Macipe et al. (2015).

The fact that the present study’s previously mentioned findings are not in agreement with others studies might be due to the number of animals used. Due to logistical constraints, it was not possible to use more than 4 animals in the present study. This sample size is much smaller than the eight animals used by Macri & Patterson-Kane (2011), the sixteen used by Cozzi et al., (2010), the eighteen used by Spielman (2000) and Martínez-Macipe et al. (2015), the thirty-four used by Frank & Houpt (1999) and the sixty-eight used by Pageat & Tessier (1997).

Additionally, personalities’ differences have a larger effect when sample sizes are small, which is in line with Gaultier et al. (2005) findings for tigers. Even within the control and the test group of this study, obvious personality differences were found. Drago was the oldest male in this study and the most laid back. He spent most of the sessions laying down on the observational platform, and he did not seem too bothered by weather changes, other lynxes nearby or human presence. On the other hand, Foco was one of the youngest and he was more alert. Despite not seeming too bothered by the presence of other lynxes nearby, he seemed more disturbed by weather changes and human presence than Drago. He was the one that vocalised the most. The test animals, Enebro and Fresco, were more restless than the animals in the control group, but showed some differences between them as well. Enebro showed more avoidance towards humans and was less interested in other lynxes than Fresco. Fresco was more temperamental whenever the keepers were nearby, and he was the only one during this study to take part in a lynx-lynx interaction (through the fence) with his cub. These few personality differences may help to explain why the results of this study are not always in line with those of the other studies.

The only significant difference found in the Pre- and Post-Experimental Study analysis was for the effects of period on the Stereotypic behaviours. In fact, the control group performed these behaviours in 5.66% and 11.44% of the scans in the three months prior to and after the study, respectively, and the test group in 7.33% and 16.72% of the scans prior to and after the study, respectively. These findings are in accordance to the ones by Spielman (2000) that showed stereotypies to increase after the application of the pheromone had ceased. Despite this, Spielman states that such a change could be due to weather changes. The fact that, in the present study, the test group showed a significant increase in Stereotypic behaviours during the post-treatment period could lead one to believe that it might be caused by the pheromone. However, the fact that the control group also showed a significant increase, suggests that it might rather be due to external factors, e.g. the relocation of the males to different enclosures, and the fact that they were paired with females for the beginning of the mating season.

As for the remaining behavioural categories, the fact that no significant differences were found in their frequency before and after this study, and between both groups, might be due to several factors. The time of the year plays an important role in the lynxes’ behaviour. The pre-experimental period, July, August and September, is the hottest time in Portugal, and most of southern Europe. Penabad et al. (2012) stated that these months are the ones when lynxes are less active, mainly because of the high temperatures. In contrast, November, March and April are the months with the highest activity levels. Also, the lynxes’ placement within the enclosure area of the CNRLI changed between the pre- and the post-experimental period. This was a consequence of the new male-female pairings for the breeding season that, as usual, starts in December. In detail, during the pre-experimental period the males used in this study were placed in the same enclosures as during the experimental period, but had the whole area of the enclosures for themselves. During the post-experimental period, the males were placed in different enclosures and shared the enclosures with females. This sharing was done gradually, as first they were separated by fences. Males and females were only put together after there were signs that the females were receptive, so that courting and copulation could take place. It is inevitable and understandable that during this period the males’ behaviour was affected by the females, since oestrus peaks between December and January (Vargas et al., 2009).

Even though several authors (Pageat & Gaultier, 2003; Lansdberg, 2006; Vargas et al., 2009) agree that pheromone therapy is an advantageous treatment against abnormal behaviours in animals, it has its limitations. Hargrave (2014) believes that pheromones alone are not able to reverse long-term, learnt behaviours, and some stereotypies fall into this category. Pheromones represent a small part in the sensory system, seeing that animals resort to other senses when assessing their environment (Hargrave, 2014). Pageat & Gaultier (2003) agree and add that in a natural setting, when animals release pheromones, they usually adopt a specific pose and may expose body parts that are normally covered. In addition, animals may modify their surroundings, when scratching or spraying certain surfaces (Pageat & Gaultier, 2003). All these actions put together help the receiving animal to detect the pheromone, by opening the VNO (Pageat & Gaultier, 2003). These additional stimuli cannot be replicated in a study like the present one. To compensate for that fact, and to ensure that the receiving animal would sense the pheromone, a larger quantity of the synthetic analogue was used (6mL per location) than what would be naturally expelled by an animal (Pageat & Gaultier, 2003).

When working with animals, especially wild species, potential obstacles that may arise every now and then must be taken into consideration. For that reason, one has to be flexible and open to change. This study was no different, and suffered from some adjustments that were not initially planned.

Drago’s and Fresco’s indoor areas (EP in Figure 2) had no cameras and the video recording software did not allow video recordings of Enebro’s and Foco’s indoor areas’ cameras. For this reason, if Drago and/or Fresco were inside the building, there was no way of knowing what they were doing there, thus they were then recorded as “fv” (out of sight). If the other two animals, Enebro and Foco, were inside their indoor areas, the observers took real-time notes of their behaviours, despite not having the ability of making video recordings. The animals used their indoor areas only a few times, especially when the weather was rainy. It was evident that rain influenced the behaviour of the lynxes used in this study, which corroborates the findings of Schmidt (1999) and Penabad et al. (2012). These authors reported that lynxes display very low activity levels during rain, which goes in line with the behaviour of the males used in this study that spent most of the rainy days sheltered and/or hiding.

One of the two main purposes of CNRLI is the successful reintroduction of Iberian lynx specimens in the wild, in order to increase this animal’s wild population (Cunha Serra et al., 2005). For that reason, the priority of the centre is the cubs and the successful completion of each one’s reintroduction training. This study was conducted at the same time as the reintroduction training was taking place, with the mother and the cubs in the main enclosure area. Thus, the males used in the study spent the entire time of this study in the husbandry area (Figure 2). Every other day, females and offspring would be moved between enclosures in the main enclosure area. On the days that females and cubs were not present in the males’ enclosure, keepers would place live prey (rabbits) in the main enclosure area, and the rabbits remained there for about a day, in plain sight of the males. This was done so that cubs could train their hunting skills when moved into that area, the following day. Schmidt (1999) found that the activity pattern of wild Eurasian lynx (Lynx lynx - Linnaeus, 1758) was mainly determined by the search and consumption of prey, and Penabad et al. (2012) found that captive Iberian lynxes’ activity pattern mirrors that of their wild conspecifics. Thus, one can extrapolate that the presence of rabbits might have had an effect on the results, as the males were “disturbed” every day, either by other lynxes or by live prey that they could not hunt.

The results of the present study are in accordance, but also contradict the ones of other studies, depending on the analysed behaviours. The small sample size, the dissimilar circumstances prior to, during and after this study, and the different personalities displayed by these animals make it difficult to draw definitive conclusions. Further studies are needed to corroborate these findings. It is my belief that a similar study done with a bigger sample, testing different synthetic analogues of feline pheromones, and in a time frame that does not overlap with the reintroduction training of the lynxes, will be more conclusive. With further research, it might even be possible to isolate a lynx-specific analogue pheromone that would be better targeted for such a study.